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LAMP primer group, kit and method for detecting capsule histoplasma capsulatum

A histoplasma and primer set technology, applied in the biological field, can solve problems that have never been reported, and achieve accurate detection, real-time detection, rapid and effective identification

Inactive Publication Date: 2021-07-23
PEKING UNION MEDICAL COLLEGE HOSPITAL CHINESE ACAD OF MEDICAL SCI +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0008] At present, the products and methods for the detection of Histoplasma capsulata by molecular means in this field are still blank, and the LAMP primer set and kit for detecting Histoplasma capsulata with high sensitivity and high specificity have never been reported in this field.

Method used

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  • LAMP primer group, kit and method for detecting capsule histoplasma capsulatum
  • LAMP primer group, kit and method for detecting capsule histoplasma capsulatum
  • LAMP primer group, kit and method for detecting capsule histoplasma capsulatum

Examples

Experimental program
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Effect test

experiment example 1

[0155] Experimental example 1 establishment of detection system

[0156] 1. Preparation of isothermal amplification reagents

[0157] The constant temperature amplification reagent includes: 1 μL of 8U / μL Bst polymerase; 0.2 μL of 1M KCL; 0.4 μL of 0.4MMgSO 4 0.2 μL of 10% Tween 20; 1.12 μL of 25 mM dNTPs; 5.52 μL of sterile water; 0.16 μL of 50 mM dye.

[0158] 2. Selection of fluorescent dyes

[0159] The fluorescent dye is Evagreen.

experiment example 2

[0160] Experimental Example 2 Establishment of constant temperature detection primers

[0161] 1. Design of LAMP primers

[0162] According to the gene sequence of Histoplasma capsulata included in GenBank, they were collected separately. Screen specific conserved regions, determine sequence status, design 10 pairs of 60 primers, compare and screen.

[0163] The primer sequences used were synthesized by China Sangong Company. After screening, 6 primers designed for the detection of the Histoplasma capsulatum gene for the target sequence SEQ ID NO: 7, including two inner primers (FIP and BIP) and two outer primers (F3 and B3) and two loops Primers (LF and LB). The nucleotide sequences of the obtained primer sets are listed in Table 1.

[0164] The specific preparation of the constant temperature amplification detection system is: 0.12 μL each of 0.3 μM outer primer F3 and outer primer B3; 0.96 μL each of 2.4 μM inner primer FIP and inner primer BIP; 0.4 μL each of 1 μM loop...

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Abstract

The invention relates to a LAMP primer group, kit and method for detecting capsule histoplasma capsulatum, and belongs to the technical field of biology. The LAMP primer group for detecting capsule histoplasma capsulatum comprises the following primers: an outer primer Hca-F3: 5'-CGATTTTGCTGCATGGATC-3', an outer primer Hca-B3: 5'-CCAATTCAGCAGCATGGTTG-3', an inner primer Hca-FIP: 5'-ACGTTTTGCTGGCCCAACTCAGAAAGGCGTTGGTACCGA-3', an inner primer Hca-BIP: 5'-TGGTTATGCGATGGAATTCGCGTCTCTGGTTGCTTCTTCGG-3', a loop primer Hca-LF: 5'-GCCAGGGGATCTAAAGAG-3', and a loop primer Hca-LB: 5'-CGAGTACCACAATCTATACC-3'. The primer group, the kit and the method provided by the invention can be used for accurately, fast and efficiently identify the capsule histoplasma capsulatum.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a LAMP primer set, a kit and a method for detecting histoplasma capsulatus. Background technique [0002] Histoplasma capsulatum is a biphasic fungus that exists in nature. It mainly invades the lung and mononuclear-phagocyte system. The hilar lymphatic system enters the blood and spreads to various organs of the whole body, forming a serious systemic disseminated disease. Histoplasmosis is a disease caused by the infection of Histoplasma capsulatum. The pathogens that cause human infection include Capsule varietal (American type) and Dubo varietal (African type), which are found in South America and Africa. Infection in my country is regionally distributed, mainly in nine provinces in the Yangtze River Basin, and the infection rate is higher in rainy and humid central and southern China, eastern China and southwest China. The causes of human histoplasma infection are mo...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6895C12Q1/6844C12Q1/04C12N15/11C12R1/645
CPCC12Q1/6895C12Q1/6844C12Q2531/119C12Q2563/107
Inventor 杨启文贾沛瑶朱盈徐英春刘莹莹韩春燕
Owner PEKING UNION MEDICAL COLLEGE HOSPITAL CHINESE ACAD OF MEDICAL SCI
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